The goal of this proposal is to request a high-throughput imaging and analysis system to support innovative research in imaged-based research for investigators at the University of Colorado, Boulder (CU-Boulder). This system is a fully integrated hardware and software system for acquisition and analysis of images for high-throughput screens of both fixed and live cells. The system includes an automated image acquisition using both laser- and image-based autofocus technology and an integrated data storage and analysis software including a variety of image processing modules, which enables this system to be used in a wide range of image-based biological assays. It also incorporates environmental control and liquid handling modules that are required for real-time live-cell imaging studies. The successful acquisition of this state-of-the-art imaging and analysis system will promote a number of collaborations between NIH- funded investigators in many disciplines. Four research projects screen for novel proteins in TGF-beta signaling, cell polarity complex formation, mitochondria metal homeostasis and p53 apoptotic responses using RNAi screen;five projects screen for novel small-molecule bioprobes, such as E3 ubiquitin ligase inhibitors, histone lysine demethylase modulators, and inhibitors of bacterial pathogenesis;three research projects focus on the real-time live-cell imaging studies including W-RAMP complex formation, TGF-beta induced Smad translocation, pain facilitation via neuron-to-glia signaling, exploring the potential of glia for regulating clinically relevant opioid actions, and role of high mobility group box 1 protein in central nervous system phenomena. Successful execution of these research projects will lead to discovery of novel therapeutic targets and lead compounds for clinical development. In addition to facilitate the proposed research projects, we also expect this system to provide a useful experience base for extending cutting-edge capabilities in high-throughput image-based screen and real-time live-cell imaging studies to the larger research community. Finally, the proposed instrument and datasets will be available for upper level graduate and undergraduate research projects through undergraduate research projects through instrument laboratory courses, a graduate chemical biology/drug discovery course, and a biophysics course. PUBLIC HEALTH RELEVANCE: The requested state-of-the-art imaging and analysis system will facilitate a number of NIH-funded research projects, such as bacterial pathogenesis, tumor suppressor p53 signaling, and pain facilitation. Successful execution of these research projects will lead to discovery of novel therapeutic targets and lead compounds for clinical development.